The present invention is directed to an improved method for synthesizing 2-hydroxy-3,17xcex2-estradiol derivatives and new intermediate compounds useful in synthesizing such estradiol derivatives.
Originally discovered as an in vivo metabolite of 3,17xcex2-estradiol, 2-methoxyestradiol has recently been shown to strongly inhibit vascularization of solid tumors making it a potential anticancer agent. Other 2-alkoxy-estradiol derivatives also possess potent anti-angiogenesis properties. A critical intermediate in the synthetic production of these 2-alkoxy-estradiol derivatives is 3,17xcex2-protected 2-hydroxy-estradiol. The family of biologically active 2-alkoxy analogs are most commonly produced from alkylation of the corresponding 3,17xcex2-protected 2-hydroxy analog.
A number of synthetic procedures exist for the production of 2-hydroxyestradiol and its subsequent conversion to the 2-alkoxy analogs. The most popular approach has been to prepare the 2-formyl-estradiol analog, treat it with an organic peracid under Baeyer-Villiger conditions and hydrolyze the resulting formate ester to the 2-hydroxy derivative. Although affording reasonable yields, these multi-step schemes are technically involved and require chromatographic purification at one or more steps. Other more direct approaches such as direct oxygenation of 2-lithiated estradiol analogs suffer from low yields and laborious purifications. Direct introduction of the 2-methoxy moiety via organometallic activation of the estradiol aromatic A-ring has been achieved. However, the difficult preparation and cost of the organometallic reagent make this synthesis unsuitable for large-scale use.
Given the anti-cancer potential of 2-alkoxy-estradiol derivatives, a need exists for a practical, scalable, high yielding and direct synthetic procedure for manufacture of such compounds.
The present invention is directed to an improved method for synthesizing 2-hydroxy-3,17xcex2-estradiol derivatives and to intermediate compounds that are useful to produce such estradiol compounds. In particular, the present invention is directed to the preparation of 2-boronyl or 2-silyl modified estradiol analogs.
In accordance with the present invention, 3,17xcex2-estradiol is reacted with a suitable organolithium reagent and either a suitable boron reagent or a suitable silicon reagent to form a compound represented by one of the corresponding structural formulae, respectively: 
(If a boron reagent) (If a silicon reagent)
where R1 and R2 can be the same or different and each represents an alkaline stable hydroxy protecting group. Preferably, R1 and R2 are each methoxymethyl. R3, R4 and R5 are selected from the group consisting of halogens, alkyl, aryl, hydroxy, substituted or unsubstituted alkoxy, and substituted or unsubstituted aryl.
The organolithium reagent is preferably sec-butyllithium. The silicon and boron reagents are represented by the following corresponding formulae: 
(silicon reagent) and (boron reagent)
R3, R4, R5, and R6 are selected from the group consisting of halogens, alkyl, aryl, hydroxy, substituted or unsubstituted alkoxy, and substituted or unsubstituted aryl. Preferably, the silicon reagent is either dichlorodimethylsilane or diethoxydimethylsilane. Trimethyl borate is the preferred boron reagent.
The boronyl or silyl modified estradiol is subsequently oxidized to form the compound represented by the following structural formula: 
where R1 and R2 are as described above. Sodium perborate is the preferred oxidizing agent for the boronyl derivative. Hydrogen peroxide is the preferred oxidizing agent for the silyl derivative.
The present invention is further directed to novel 2-boronyl or 2-silyl modified estradiols and methods of preparing these novel compounds, which are useful intermediates in forming 2-hydroxy-3,17xcex2-estradiol derivatives.
In accordance with the present invention, methods of synthesizing the foregoing novel compounds are provided. 2-alkoxy estradiols and a numbering system for identifying each carbon atom in the estradiol ring system are shown in the following structural formula: 
where R10 is an alkyl group or a substituted alkyl group.
As used in the description, and in the claims, xe2x80x9calkylxe2x80x9d represents a straight-chain or branched hydrocarbon radical of 1 to 10 carbons in all its isomeric forms, such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl, etc. The alkyl group can optionally contain one or more double or triple bonds. The term xe2x80x9carylxe2x80x9d signifies a phenyl-, or an alkyl-, nitro- or halo-substituted phenyl group. The term xe2x80x9calkoxyxe2x80x9d signifies the group alkyl-Oxe2x80x94. Suitable substituents on an alkyl group or phenyl group include one or more halogens (e.g., fluoro, chloro, bromo and iodo), nitro, nitrile, xe2x80x94NH2, xe2x80x94NH (alkyl), xe2x80x94NH (substituted alkyl), xe2x80x94N (alkyl)2, xe2x80x94N(substituted alkyl)2, carbonyl groups, xe2x80x94CONH2, xe2x80x94CONH(alkyl), xe2x80x94CONH(substituted alkyl), CON(alkyl)2, xe2x80x94CON(substituted alkyl)2, xe2x80x94CO2H, xe2x80x94COO(alkyl) and xe2x80x94COO (substituted alkyl). Halogenated alkyl groups can contain more than one kind of halogen. Examples of suitable alkyl or substituted alkyl groups include methyl, ethyl, n-propyl, iso-propyl, trifluoromethyl, trifluoroethyl, NO2xe2x80x94CH2xe2x80x94CH2xe2x80x94, (CH3)Nxe2x80x94CH2xe2x80x94CH2xe2x80x94, xe2x80x94CH2CH2xe2x80x94COxe2x80x94Rxe2x80x2, wherein R10 is xe2x80x94H, alkyl, substituted alkyl, xe2x80x94OH, xe2x80x94O (alkyl), xe2x80x94O(substituted alkyl), xe2x80x94NH2, xe2x80x94NH(alkyl), xe2x80x94NH (substituted alkyl), xe2x80x94N(substituted alkyl)2 and xe2x80x94N(alkyl)2.
The following reaction scheme depicts the production of 2-methoxyestra-3,17xcex2-diol (5) by preparing a 2-boronyl or 2-silyl modified estradiol analog in accordance with the present invention. This synthesis is described in greater detail below.

In accordance with the foregoing reaction scheme, a first intermediate (2) is prepared from 3,17xcex2-estradiol (1) by protecting the two hydroxyl groups to form a compound represented by Structural Formula (I): 
where R1 and R2 may be the same or different and each is independently a hydroxyl protecting group. As used herein, hydroxyl refers to alcohols and phenolic groups. A suitable xe2x80x9cprotecting groupxe2x80x9d is substantially inert with respect to the reagents used in the subsequent reactions in the disclosed synthesis of 2-alkoxy estradiols and does not cause, for example, undesired side reactions. Hydroxyl protecting groups are well known in the art and are described in, for example, Chapters 2 and 3 of Greene and Wuts, xe2x80x9cProtective Groups in Organic Synthesisxe2x80x9d, John Wiley and Sons (1999), the entire disclosure of which is herein incorporated by reference. The skilled artisan can select suitable groups for use in the disclosed synthesis as well as conditions for applying and removing the hydroxyl protecting groups.
Preferably, R1 is a protecting group that promotes ortho-lithiation at the 2-position. Such a protecting group contains an atom bearing a lone pair of electrons xcex2 to the hydroxyl oxygen. Such an atom is a heteroatom and preferably an oxygen atom. The general description of these types of ortho-directing protecting groups are substituted methyl ethers such as methoxymethyl, 2-methoxyethoxymethyl, 2-(trimethylsilyl)ethoxymethyl, tetrahydropyranyl, and tetrahydrofuranyl. Methoxymethyl is most preferred for its selective promotion of lithiation at the 2-position over the 4-position. Correspondingly, it is preferred that R1 and R2 are each methoxymethyl.
The first intermediate (2) is reacted with an organolithium reagent and a boron or a silicon reagent to form a second intermediate having Structural Formula (II) from the boron reagent or (III) from the silicon reagent: 
where R1 and R2 are as described above with respect to Structural Formula (I) and R3, R4 and R5 are halogens, alkyl, aryl, hydroxy, substituted or unsubstituted alkoxy, or substituted or unsubstituted aryl.
Organolithium reagents are well known in the art. Examples of suitable organolithium reagents include methyllithium, n-butyllithium, sec-butyllithium, tert-butylithium, or phenyllithium. Preferably, the organolithium reagent is sec-butylithium. Reacting the first intermediate (2) with a 1 to 3 fold mole excess of organolithium reagent is preferred. A 1.5 fold mole excess of sec-butyllithium is most preferred.
The silicon and boron reagents are represented by Structural Formulae (IV) and (V) respectively: 
where R3, R4, R5, and R6 are halogens, alkyl, aryl, hydroxy, substituted or unsubstituted alkoxy, or substituted or unsubstituted aryl. The preferred silicon reagent is either dichlorodimethylsilane or diethoxydimethylsilane. Suitable boron reagents are tributyl borate, tripropyl borate, triisopropyl borate, triethyl borate, and trimethyl borate. The preferred boron reagent is trimethyl borate. Preferably, the first intermediate (2) is reacted with a 1 to 5 fold mole excess of either the silicon or boron reagent.
Suitable solvents for the organolithium and boron or silicon reactions include etheral solvents such as tetrahydrofuran (THF), dioxane, or glyme. THF is the preferred solvent. Suitable temperatures for carrying out the reaction typically are in the range of from about xe2x88x92110xc2x0 C. to about 0xc2x0 C. A reaction temperature below xe2x88x9270xc2x0 C. is preferred.
The second intermediate is reacted with an oxidizing agent to form a third intermediate (3) having the Structural Formula (VI): 
where R1 and R2 are as described above with respect to Structured Formula (I). Suitable oxidizing agents include peracetic acid, 3-chloroperoxybenzoic acid, tert-butyl hydroperoxide, magnesium monoperoxyphthalate, potassium peroxymonosulfate sold under the trademark OXONE(copyright), hydrogen peroxide and sodium perborate. Sodium perborate is the preferred oxidizing agent.
A suitable biphasic solvent system for the oxidation reaction includes etheral solvents, such as THF, dioxane, or glyme, and water. THF and water is the preferred solvent system. A suitable THF-to-water ratio for the solvent system ranges from about 1:1 to about 1:5. Preferably, a 1:2 ratio of THF to water is used. A suitable temperature range for performing the oxidation reaction is between about 10xc2x0 C. to about 35xc2x0 C. Preferably, the oxidation reaction is performed at about room temperature.
Prior to oxidation, the boronyl (II) or silyl (III) second intermediate may be isolated. For example, quenching the borane reaction hydrolyzes the dialkoxy boronyl intermediate to form the boronic acid estradiol analog. Specifically, the boronyl modified estradiol analog represented by Structural Formula (VII) has been isolated: 
where R1 and R2 are each methoxymethyl and R3 and R4 are hydroxy groups. Specific conditions for quenching the borane reaction are provided in Example 3. Other modified estradiols include, but are not limited to, 2-trimethylsilyl, 2-chlorodimethylsilyl and 2-dimethylethoxysilyl. Preferably, the boronyl (II) or silyl (III) second intermediate remains in solution and is oxidized directly to form the third intermediate (3).
The 2-hydroxy group of the third intermediate (3) is xe2x80x9calkylatedxe2x80x9d to form a fourth intermediate (4) represented by Structural Formula (VIII): 
where R1 and R2 are as described above with respect to Structured Formula (I) and R10 is a substituted or unsubstituted alkyl group. Preferably, R10 is a methyl group. As used herein, xe2x80x9calkylationxe2x80x9d refers to the removal of the phenolic proton under basic conditions and subsequent reaction of the phenoxide anion with an alkylating agent. The alkylation of phenols is well known in the art and can be accomplished by the xe2x80x9cWilliamson Reaction,xe2x80x9d for example. Specific conditions for alkylation are set forth in Example 3.
The hydroxyl protecting groups at the 3 and 17 positions of the fourth intermediate (4) are deprotected to form the desired 2-alkoxy estradiol represented by Structural Formula (IX): 
R10 is as described above. Suitable conditions for the removal of phenolic protecting groups are commonly known in the art and are disclosed in Chapters 2 and 3 of Green and Wuts, xe2x80x9cProtective Groups in Organic Synthesisxe2x80x9d, John Wiley and Sons (1999). Specific conditions for deprotection are provided in Example 4.